Molecular monitoring of tumor cell contamination in leukapheresis products from stage IV neuroblastoma patients before and after positive CD34 selection.
BACKGROUND
Autologous peripheral blood stem cell (PBSCs) are frequently used to reconstitute hematopoiesis following administration of megatherapy in children with advanced stage IV neuroblastoma. Some centers prefer the use of autografts enriched for CD34+ progenitor cells because the positive selection procedure is believed to reduce indirectly tumor cell contamination.


METHODS
In this study, we monitored the efficiency of tumor cell purging following CD34 selection in PBSCs from seven patients with advanced neuroblastoma by using a highly sensitive reverse transcriptase-polymerase chain reaction (RT-PCR) analysis, Amplification of tissues-specific mRNA transcript of tyrosine hydroxylase gene with nested primers enabled the detection of residual neuroblastoma cells with a sensitivity of one malignant-cell per 10(6) normals.


RESULTS
Using this method, contaminating tumor cells were detected in seven of nine leukapheresis products of the patients. After positive immunoselection of CD34+ cells on Ceprate column, only one of nine enriched stem cell fraction still contained tumor cells detectable by RT-PCR. In six cases, PCR positive PBSCs became PCR negative after selection.


CONCLUSIONS
We conclude that tumor cell contamination may be frequently detected in PBSC harvests of stage IV neuroblastoma patients by sensitive molecular analysis. The load of contaminating malignant cells might be reduced following CD34 selection.
Tchirkov
A
A
Unité Bioclinique de Prélèvement et de Traitement des Cellules Souches Hématopoïétiques et Secteur d'Etude en Hématopoïése, Service de Pédiatrie B. France.
Kanold
J
J
Giollant
M
M
Halle-Haus
P
P
Berger
M
M
Rapatel
C
C
Lutz
P
P
Bergeron
C
C
Plantaz
D
D
Vannier
J P
JP
Stephan
J L
JL
Favrot
M
M
Bordigoni
P
P
Malet
P
P
Briançon
G
G
Deméocq
F
F
D018952:Antigens, CD34 / Q000032:analysis*; D002675:Child, Preschool; D005260:Female; D018380:Hematopoietic Stem Cell Transplantation; D006801:Humans; D018189:Immunomagnetic Separation / Q000592:standards; D007223:Infant; D007937:Leukapheresis; D008297:Male; D009360:Neoplastic Cells, Circulating; D009447:Neuroblastoma / Q000276:immunology / Q000473:pathology / Q000628:therapy*; D016133:Polymerase Chain Reaction; D012333:RNA, Messenger / Q000032:analysis; D014446:Tyrosine 3-Monooxygenase / Q000235:genetics
D018952:Antigens, CD34; D012333:RNA, Messenger; D014446:Tyrosine 3-Monooxygenase